Carboxymethyl guar gum silver nanocomposite film: Preparation and antimicrobial activity

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1 Carboxymethyl guar gum silver nanocomposite film: Preparation and antimicrobial activity DEVENDRA KUMAR VERMA* and ANEK PAL GUPTA Department of Applied Chemistry and Polymer Technology, Delhi Technological University (Formerly Delhi College of Engineering), Shahabad, Daulatpur, Bawana Road, Delhi , India ABSTRACT Silver nanoparticles have been utilized in various applications because of its highly conductive and antimicrobial properties. A new method for the synthesis of silver nanoparticles has been used by reduction process to prepare carboxymethyl guar gum- silver (CMGG/Ag) nanocomposite as an antimicrobial film for food packaging applications. The nanoparticles and composites are tested by XRD, FTIR, SEM, TEM, EDX, UV and UTM. Antimicrobial property is tested by using gram positive and gram negative bacteria. Key Words: Carboxymethyl guar gum, silver nanoparticles, nanocomposite film, antimicrobial activity, food packaging INTRODUCTION A huge commercial demand of nanoparticles is for wide applicability in various areas such as electronics, catalysis, chemistry, energy, and medicine 1. Especially silver nanoparticles (Ag NPs) are being interested among scientists because of their different antibacterial, catalytic and specific physical and optical properties 2. There have been a lot of reports for synthesis of Ag NPs, for example, electrochemical techniques 3, photochemical reactions in reverse micelles 4, chemical reduction 5 and green synthesis 6. The reliable, easy, fast, and eco-friendly method is till awaited for preparation of highly pure Ag NPs. In this article glycerol as green reducing agent and sodium hydroxide as catalyst has been used for preparation Ag NPs. In recent years, antimicrobial packaging has been found one of the most reliable and promising tool for the next generation of active packaging 7. It is well known that silver ions and silver-based compounds are highly toxic to microorganisms showing strong biocidal effects on as many as 16 species of bacteria including E. coli. Silver, as an antibacterial component, have been used in the formulation of dental resin composites, ion exchange fibres and in coatings of medical devices 8. Undoubtedly, the most prominent area of research and development is food packaging. The biggest benefit of silver as antimicrobials is that silver can be easily mixed into different materials such as plastics and textiles for desirable uses, where as traditional antimicrobials would be impractical. Ag NPs make nanocomposites very comfortably with nonbiodegradable plastic or synthetic polymers. But this is also fact that these synthetic non biodegradable polymers have created a tough situation for the living being for a healthy life 9. This scenario prompted us to change the direction of our research from nonbiodegradable to the biodegradable polymeric products. In this article we have introduced carboxymethyl guar gum (CMGG) as a matrix for CMGG/Ag nanocomposite. Guar and its derivatives are all biodegradable and have been used in many applications including food, drugdelivery and health care products because of their natural abundance and their low cost and other desirable functionalities 10. CMGG is an anionic semi-synthetic guar gum derivative 11,12. CMGG is a cheap and easily water soluble commercial polysaccharide because it covers a wide range of industrial applications, and has a good film forming property 13. In this communication, we have reported the synthesis and characterization of Ag NPs by a reduction process and then preparation of CCMG/Ag nanocomposites by ex-situ method to study the antibacterial activity for food packaging applications. EXPERIMENTAL Materials CMGG powder has been obtained from Hariom gum industry, Gujarat, India and used without further purification. Sodium hydroxide, silver nitrate (AgNO 3 ) and glycerol have been purchased from Merck, Germany. All aqueous solutions were made using ultrahigh deionized (DI) water. Preparations of Ag nanoparticles (Ag NPs) Ag NPs were synthesized by reduction process by using glycerol as a reducing agent and sodium hydroxide as a catalyst. Many researchers have used the glycerol as reducing agent 14,15. The importance thing of this process was the quick synthesis of Ag NPs and no wastage of chemicals. Glycerol is very eco-friendly and sodium

2 hydroxide can be reused. For the synthesis of Ag NPs, a stock solution of 0.2 M AgNO 3 was prepared in DI water. The solution has been mixed with 2 ml glycerol solution followed by addition of sodium hydroxide (0.5 M) drop wise at room temperature with continuous stirring. Immediately it formed yellowish black colloidal solution indicating the formation of Ag NPs. The solution was centrifuged and dried at 20 0 C and stored in the desiccator until further use. Preparation of carboxymethyl guar gum-silver nanoparticle (CMGG/Ag) nanocomposite film CMGG/Ag nanocomposites have been synthesised in one step by ex-situ method. First of all 1% w/v CMGG solution have been made using DI water. For this 1 gm of CMGG powder was mixed in 100 ml of DI water in 250 ml beaker with constant stirring for 30 min, requisite amount of Ag NPs (1% w/w of CMGG) have been mixed with this solution with constant stirring for 4-5 h until we get clear viscous solution of CMGG/Ag. Then CMGG/Ag nanocomposite film was prepared by solution casting method. This nanocomposite film was dried in room temperature for overnight at room temperature. Characterization of synthesized AgNPs and CMGG/Ag nanocomposite film The size, morphology and composition of the nanoparticles and nanocomposites have been tested by of scanning electron microscope (SEM) (HITACHI-S- 3700), transmission electron microscopy (TEM), energy dispersive X-ray analysis (EDX) (BRUKER D8- ADVANCE, Fourier Transform infrared spectroscopy (FTIR) (Thermo scientific Nicolet 380) and ultravioletvisible spectrophotometer (UV- Vis, Parkin Elmer Lambda -750). The tensile strength of the films was tested with UTM ( Universal Testing Machine ) and compared with standard biodegradable corn packaging films. CMGG/Ag nanocomposite films have also been tested against antimicrobial activity against gram positive (S. Bacillus) and gram negative (E. coli) bacteria, for different Ag NPs concentration with variation of CMGG. Antimicrobial testing was analysed by Kirby Bauer method by using Luria Broth as a bacteria growing media. RESULTS AND DISCUSSION UV Vis, SEM and TEM analysis of Ag NPs The prepared Ag NPs shows an intensive UV vis absorbance band around 411 nm. In general, the Ag NPs often shows an intense SPR band 16 from 350 to 450 nm (Szunerits & Boukherroub, 2006). According to the reported literature, the position of UV-absorption of Ag NPs depends on their size and shape as well as their surrounding medium 17. Thus it clarifies that the particles are not homogeneous in size, but there is of great change in size. The morphologies of the prepared Ag NPs and CMGG/Ag nanocomposite film are done by using SEM and TEM (Fig. 1(a), (b), (c), (d), (e) and (f)). From the SEM images (Fig.1a and 1b), it is cleared that nanoparticles have been formed in a spherical shape. Further the TEM images of the nanoparticles (Fig.1c and 1d) shows clear formation of Ag NPs in range 20 nm to 70 nm in agglomeration form. The average particle size was found ~ 40 nm. From Fig. 1e and 1f it can be seen clearly that Ag NPs are well dispersed in the CMGG matrix. This confirms the formation of CMGG/Ag nanocomposite film. 1a) 1b)

3 1c 1d) 1e) 1f) Fig.1. (a) and (b) SEM images of spherical silver nanopariticles, (c) and (d) TEM images of Spherical silver nanoparticles showing different sizes of spherical nanoparticles, (e) and (f) SEM images of CMGG/Ag nanocomposite films. Fourier Transform Infrared Spectroscopy (FTIR) and EDX analysis The infrared spectra of silver nanoparticles, CMGG and CMGG/Ag nanocomposite film are shown in Fig. 2 (a), (b), (c). No peak was obtained in Ag NPs FTIR spectrum, it shows that silver nanoparticles were pure and, no chemical bond was attached to Ag metal during the synthesis of Ag NPs. Formation of silver nanoparticle was also proved by EDX analysis (Fig. 2d). The characteristic broad band for O-H group of CMGG appeared at 3438 cm -1. A peak around 2928 cm -1 was attributed to an asymmetrically stretching vibration of C- H band in alkanes. The asymmetrical and symmetrical vibrations due to moiety were assigned to 1604 cm -1 and 1422 cm -1 which may be attributed to carboxymethyl groups into guar gum molecule. Several absorption bands between ( ) cm -1 were attributed to the contribution of various functional groups, such as C-O and C-O-C bondings. All these peaks were found in pure CMGG powder 18. The spectral band for CMGG/Ag nanocomposite film (Fig. 2c) appears with slide shifting of peaks for O-H group, 2928 cm -1 to 2918 cm -1 for asymmetrically stretching vibration of C-H band in alkanes, 1604 cm -1 and 1422 to 1637 cm -1 and 1430 cm -1 for carboxymethyl groups in to guar gum molecule. This shows that the there was no covalent formation between Ag NPs and CMGG.

4 (d) Fig.2. FTIR spectrum of silver nanoparticles synthesized by reduction process (a), FTIR pattern of CMGG powder (b),ftir spectrum of CMGG/Ag nanocomposite synthesized by exsitu method (c) and EDX patter of the silver nanoparticles (d). XRD analysis The nature of the silver nanoparticles synthesized by such process has been evaluated in free stage as well as in CMGG/Ag nanocomposite by XRD method. The XRD of CMGG and Ag NPs was compared. XRD of CMGG shows amorphous nature (Fig. 3b), while the XRD profile of Ag nanoparticles in free state, as well as in composite state, presented in Fig. 3.(a) and (c) exhibit characteristic peaks at scattering angle (2θ) 38.12, 44.46, 64.5, (Fig. 3a) and 38.14, 44.36, 64.56, (Fig. 3c), corresponding with scattering from the (111), (200), (220), and (311) planes, respectively. These diffraction peaks firmly represent a face centred cubic structure of crystalline silver nanoparticles 19,20. This also represents a high purity of synthesized silver crystal.

5 Fig.3. XRD pattern of silver nanoparticles( face centred cubic lattice)synthesized by green synthesis(a), XRD spe ctrum of CMGG powder(b), XRD spectrum of CMGG/Ag nanocomposite film prepared by solution casting method(c). Antimicrobial activity The antimicrobial susceptibility of CMGG/Ag nanocomposite film is evaluated by using the disc diffusion of Kirby-Bauer method. This method has been recommended by the national committee for clinical laboratory standards (NCCLS)(NCCLS,03). CMGG/Ag nanocomposite films of different concentration (A1C = 0.5 wt %, A2C =0.75 wt %, A3C =1.0 wt %, A4C =1.25 wt %, C= 0 wt % of Ag NPs of CMGG powder) were cut in the circular shape with same size and put in the inoculated plates and stored for 24 h at 37 o C in incubator. After 24 h, incubation zones of inhibition were measured. It was found that CMGG/Ag nanocomposite at a certain level inhibited bacterial growth by more than 90%. The diameters of inhibition zones (in mm) around the different CMGG- Ag nanocomposite films are shown in Table 1. Table 1 CMGG/Ag nanocomposite films with variable amount of silver nanoparticles (wt %) and respective inhibition zones Variable amount of silver nanoparticles in CMGG/Ag nanocomposite (wt %) and respective inhibition zones in mm. Bacterial A1C film A2C film A3C film A4C film Pure CMGG film (C) Gram positive Biodegradable Gram negative Tensile strength Tensile strength of the CMGG/Ag nanocomposite films are determined by using Universal Testing Machine (UTM). Comparison study of the tensile strength is done for different concentrations of CMGG/Ag nanocomposites films with standard corn natural polymer film available in the market. For the testing of CMGG/Ag nanocomposite films, CMGG film and corn film, all the films were cut in the fine strips as mentioned in standard of UTM. Length, weight and thickness of all the strips were measured by using electronic vernier calliper by two times and their average value was used. Tensile strength of CMGG-Ag nanocomposites films of different concentrations (A1C=0.5%, A2C=0.75%, A3C=1.0 %, A4C=1.25 %,) were found respectively MPa, MPa, MPa and MPa. Tensile strength of CMGG film and corn film were found MPa and MPa respectively. Thus from tensile results it was found that as the concentration of the silver was increased from 0.5 wt% to 1.25 wt% the tensile strength also increased, which was good agreement with literature reports 21,22. The tensile strength of nanocomposites was found greater than pure CMGG film and corn films. This result explains that prepared CMGG/Ag nanocomposite film may be used as antimicrobial films for food packaging applications. CONCLUSIONS Ag NPs were synthesized successfully via reduction process using green reducing agent (glycerol). Silver nanoparticles were found of spherical shape of average size nm. Silver nanoparicles were pure and in crystalline form. Bulk amount of silver nanoparticles in a very less time can be synthesized by this method. This method was found very fast, easy and reliable with no wastage of chemicals. The CMGG/Ag nanocomposite

6 film was successfully prepared by solution casting method. A little amount of nanocomposite film was found sufficient for antimicrobial activity for both types of bacteria. Tensile strength of the nanocomposite films were also found very good and greater than of corn film and pure CMGG films. It was concluded that in future with slight improvement, CMGG/Ag nanocomposite films can be used in food packaging. REFERENCES 1. Geethalakshmi R and Sarada D V L, International Journal of Engineering Science and Technology, 2(5), 970 (2010). 2. Król-Gracz A, Nowak P, Michalak E and Dyonizy A, Proceedings of the International Congress on Advances in Applied Physics and Materials Science, Antalya Acta Physica Polonica, A 121 (2011). 3. Rodriguez-Sanchez L, Blanco M C and Lopez- Quintela M A, J. Phys. Chem., 104 (41), 9683 (2000). 4. Tamura S, Takeuchi K, Mao G, Csencsits R, Fan L, Otomo T, Saboungi M-L, Journal of Electroanalytical Chemistry, (2003). 5. Tripathi R N, Saxena A, Gupta N, Kapoor H and Singh R P, Digest Journal of Nanomaterials and Biostrucutres, 5, 2 (2010). 6. Singh C, Sharma V, Naika P K, Khandelwal V and Singh H, Digest Journal of Nanomaterials and Biostructures, 6, 535 (2011). 7. Tripathi S, Mehotra G K and Dutta P K, Bull. Mater. Sci., 34(1), 29 (2011). 8. Timothy V. Duncan, Journal of Colloid and Interface Science, 363, 1 (2011). 9. Gupta A P and Verma D K, International Journal Of Biological Macromolecule, 68, 247 (2014). 10. Sreenivasan Soumyaa R, kumar Ghoshb S, Abrahama E T, International Journal of Biological Macromolecules, 46, 267 (2010). 11. Gong H, Liu M, Chen J, Fei Han, Gao C and Zhang B, Carbohydrate Polymers, 88(3), 1015 (2012). 12. Gupta A P and Verma D K, International Journal of Advanced Research, 2 (1), 680 (2014). 13. Garg P, Studies on Hydrogels Based on Polyacrylamide and Guar Gum Derivatives, A Major-II Dissertation Submitted to Delhi Technological University India, Alba E. Díaz-Álvarez and Victorio Cadierno, Appl. Sci., 3, 55 (2013). 15. Sinha A and Sharma B P, Bull. Mater. Sci., 28(3), 213 (2005). 16. Szunerits S and Boukherroub R, Langmuir, 22, 1660 (2006). 17. Prasad B L V, Stoeva S I, Sorensen C M, Zaikovski V, Klabunde K J, J Am Chem Soc 125, (2003). 18. Dodi G, Hitcu D and Popa M I, Cellulose Chemistry and Technology, 45(3-4), 171 (2011). 19. Kirubaharan C J, Kalpana D, Lee Y S, Kim A R, Yoo D J, Nahm K S and Kumar G G, Industrial and Engineering Chemistry Research, 51, 7441 (2012). 20. Wen X, Xie Y T, Mak M W C, Cheung K Y, Li X Y, Renneberg R and Yang S, Langmuir, 22, 4836 (2006). 21. Islam Md S, Masoodi R and Rostami H 2013 Hindawi Publishing Corporation, Journal of Nanoscience, , 10 (2013). 22. Perumalraj R, ISRN Chemical Engineering, , 4 (2012) Received: 30 January 2015 Accepted: 10 March 2015

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